Effect of KLF10 deficiency on hindlimb ischemia. (A-E) WT or KLF10^−/− mice underwent femoral artery ligation to induce hindlimb ischemia. (A) KLF10^−/− mice (n = 2 of 21) developed autoamputation of the ischemic leg. Mice were photographed with a Nikon Coolpix 4600 digital camera. (B) Images (left) are representative of blood flow recovery for each time point over 28 days. Quantitation (right) of blood flow recovery was calculated as the mean blood flow (right [ischemic] leg)/left [(nonischemic] leg) by laser Doppler imaging (785-nm near-infrared Laser Doppler Imager-2; Moor Instruments).*P < .01 versus WT; **P < .05 versus WT. (C) FACS analyses of circulating PACs (Sca-1⁺/VEGFR2⁺) in WT or KLF10^−/− mice 7 days after femoral artery ligation (n = 9-10 per group). *P < .05 versus WT. (D) WT or KLF10^−/− PACs (1:1 mix of CMP- and GMP-derived PACs) were intramuscularly injected immediately after femoral artery ligation in KLF10^−/− mice. Mean blood flow recovery (ischemic leg/nonischemic leg) was measured by laser Doppler imaging after 3 days. *P < .05 versus WT +PBS. (E-F) Frozen sections of quadriceps muscles harvested 3 days after intramuscular injection of WT or KLF10^−/− PACs were labeled with cell tracker (red) and a FITC-conjugated mAb to CD31 (green). (E) Sections were analyzed for CD31 staining with the use of an AQUA/PM2000 Imaging Platform (HistoRx), and automated quantitative analysis was performed with Software suite Version 2.2 (HistoRx). *P < .01 versus WT; n = 4 mice per group. (F) Sections were examined with an Olympus, Fluoview, Model FV1000 camera at 10× magnification and FV10-ASW Version 02.01 software to determine the percentage of labeled PACs (red) that colocalized (yellow) with CD31⁺ cells (green). *P < .01 versus WT; n = 4 mice per group.